Abstract
As more and more research efforts have been attracted to dynamic or differential proteomics, a method with high temporal resolution and high throughput is required. In present study, a 35S in vivo Labeling Analysis for Dynamic Proteomics (SiLAD) was designed and tested by analyzing the dynamic proteome changes in the highly synchronized A549 cells, as well as in the rat liver 2/3 partial hepatectomy surgery. The results validated that SiLAD technique, in combination with 2-Dimensional Electrophoresis, provided a highly sensitivity method to illustrate the non-disturbed endogenous proteins dynamic changes with a good temporal resolution and high signal/noise ratio. A significant number of differential proteins can be discovered or re-categorized by this technique. Another unique feature of SiLAD is its capability of quantifying the rate of protein expression, which reflects the cellular physiological turn points more effectively. Finally, the prescribed SiLAD proteome snapshot pattern could be potentially used as an exclusive symbol for characterizing each stage in well regulated biological processes.
Highlights
To decipher the intricate cellular activities more effectively at protein level, more and more researches have been focused on dynamic proteomics [1,2,3,4,5,6] or differential proteomics [7,8,9]
A novel method to examine the dynamic proteome expression changes based on 35S metabolic pulse labeling was proposed (Figure 1). 2-Dimensional Electrophoresis (2-DE) was employed in this methodology to detect the protein changes, since it is still the best technology to directly visualize the largest number of proteins simultaneously and separately, allowing us to detect most of the differential changing proteins before the Mass Spectrometry (MS/MS) identification
There are 1651 spots matched in the two images, so the number of spot exclusively shown in phosphor-image is 613, account for 25.4% in the total 2,412 spots detected in this gel, compared with only 6.1% spots exclusively shown in the Coomassie Brilliant Blue-R 350 (CBB) staining
Summary
To decipher the intricate cellular activities more effectively at protein level, more and more researches have been focused on dynamic proteomics [1,2,3,4,5,6] or differential proteomics [7,8,9]. A novel method to examine the dynamic proteome expression changes based on 35S metabolic pulse labeling was proposed (Figure 1). The auto-radiography of the 35S short pulse labeled gel provides the information about protein newly expressed without interference from the former existed amount. Using 35S to pulse label several time points in a biological process is, for the first time, allowing us to chase the proteome expression changes at the second order derivate level
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